The estrogen receptor (ER) is a ligand-inducible transcription factor which regulates the expression of a variety of genes including some growth factors, hormones and oncogenes important for the growth of breast cancer (1, 2). Expression of the ER plays an important role in the pathogenesis and maintenance of breast cancer. In breast cancer patients, about two-thirds of tumors are ER-positive (3); 50% of these ER-positive tumors are estrogen-dependent and respond to endocrine therapy (4,5). Breast carcinomas occurring in postmenopausal women are often ER-positive (6), and many of these tumors express significantly more ER than does the normal mammary epithelium (7).
On the other hand, it has been proposed that the cumulative exposure of breast tissue to hormones associated with ovarian activity is a major determinant of breast cancer risk (8, 9). Furthermore, some environmental agents and dietary components might influence breast cancer development by functioning as xenoestrogens or estrogenic potentiating factors (10).
The ER gene spans 140 kb and is comprised of 8 exons which are spliced to yield a 6.3 kb mRNA, encoding a 595-amino acid protein with a molecular weight of 66 kilodaltons (11, 12). The ER protein is comprised of several discrete functional domains (13). Two transcriptional activation functions (TAF-1 and TAF-2) reside in exon 1 and 8 respectively (14-15). The DNA-binding domain of about 70 amino acids is located at exon 2 and 3. Exon 4 through 8 translating into 250 amino acids in size are necessary and sufficient for ligand binding (13, 16). A 22 amino acid sequence necessary for subunit dimerization has been located in exon 7 (17).
Recently, it has been reported that both the wild type and variant estrogen receptors were coexpressed in some human breast carcinoma cell lines (18-24). Variant human breast tumor ER with constitutive transcriptioanl activity has been identified in tumor specimens (25 & 26). Therefore, estrogen receptor variants might have a role in human breast cancer. However, to date, the role of the estrogen receptor variant in carcinogenesis and the regulation of its expression are poorly understood.
The human mammary gland contains a small but distinct population of ER-positive cells, comprising about 7% of the total epithelial cell population from all biopsies (27). The ER-positive cells were distributed as scattered single cells, most of them (87%) were luminal epithelial cells or occupied an intermediate position in the duct wall. The highest frequency of ER-positive cells has been found in the lobules as compared to the interlobular ducts (27). Normal HBEC grown in the commonly used media, MCDB 170 (28) and DFCI-1 (29), including the commercially available normal HBEC (Clonetics), exhibited basal epithelial, but not luminal epithelial cell characteristics. These cells have not been shown to express the estrogen receptor with significant frequency.
Recently, we have developed a culture method to grow two morphologically and antigenically distinguishable normal human breast epithelial cell (HBEC) types from reduction mammoplasty (30). Type I HBEC is deficient in gap junctional intercellular communication (GJIC) and has luminal and stem cell characteristics, i.e. the differentiation of Type I into Type II HBEC by a cyclic AMP-inducing agent (30) and the unique ability of Type I HBEC to form budding and ductal structures on MATRIGEL matrix (31), whereas Type II HBEC is capable of GJIC (30, 32-33) and expresses basal epithelial cell phenotypes (30). See U.S. ser. No. 08/308,118 filed Sep. 16, 1994 now U.S. Pat. No. 5,650,317, incorporated herein by reference.
It is an object of the present invention to describe a substantially purified human breast epithelial cell type (such as Type I HBEC) which has stem cell characteristics and which expresses human estrogen receptor.
It is also an object of the present invention to describe the expression of estrogen receptors in these two types of human breast epithelial cell (HBEC) and the neoplastically transformed cell lines derived from these cells.